Previous work on this project has defined the purification and subunit composition of brain sodium channels, reconstituted their functional properties in purified form, and described their biosynthesis and assembly in neurons. cDNAs encoding Type IIA sodium channels have been cloned and expressed in mammalian cells. Regulation of sodium channel function through phosphorylation by cAMP-dependent protein kinase and protein kinase C has been analyzed in whole cell voltage clamp and single channel recording experiments. Structure-function studies with site-directed antibodies and site-directed mutagenesis have identified regions of the channel that are critical for voltage-dependent activation, fast channel inactivation, binding of alpha-scorpion toxins, and regulation of the channel by protein phosphorylation. The proposed research aims to build on this previous work to define several related aspects of sodium channel structure and function concerning mechanisms of channel inactivation regulation by protein phosphorylation, and modulation by drugs and neurotoxins. The functional role of individual amino acid residues in a inactivation gating loop will be probed by site- directed mutagenesis, expression in mammalian cells, and patch clamp recording; the mechanism of coupling of activation to inactivation will be analyzed through mutagenesis of amino acid residues in likely coupling segments. The sites of phosphorylation of sodium channel alpha subunits by protein kinase C and cAMP-dependent protein kinase that are responsible for down regulation of channel activity and modulation of channel inactivation will be identified by antibody mapping and protein chemistry, and the functional effects of their phosphorylation will be studies by mutagenesis expression, patch clamp recording, and biochemical analysis. Receptor sites for local anesthetics and sodium channel-directed anticonvulsant drugs on the intracellular surface of the sodium channel will be probed by mutagenesis and expression methods, and the molecular mechanisms of interaction of inactivation gating with the frequency- and voltage- dependent binding of these drugs will be analyzed. Receptor sites for alpha- and beta-scorpion toxins on the extracellular surface of the sodium channel will be identified by antibody mapping and protein chemistry, and their role in the normal coupling of activation to inactivation and the pharmacological effects of the toxins on these processes will be probed by mutagenesis and expression methods. The results of these studies will give new insight into the molecular mechanism of inactivation gating of sodium channels, its coupling to voltage-dependent activation, and its modulation by protein phosphorylation, local anesthetic and anticonvulsant drugs, and scorpion neurotoxins.
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| Lenaeus, Michael J; Gamal El-Din, Tamer M; Ing, Christopher et al. (2017) Structures of closed and open states of a voltage-gated sodium channel. Proc Natl Acad Sci U S A 114:E3051-E3060 |
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| Catterall, William A (2017) Forty Years of Sodium Channels: Structure, Function, Pharmacology, and Epilepsy. Neurochem Res 42:2495-2504 |
| Catterall, William A; Wisedchaisri, Goragot; Zheng, Ning (2017) The chemical basis for electrical signaling. Nat Chem Biol 13:455-463 |
| Southan, Christopher; Sharman, Joanna L; Benson, Helen E et al. (2016) The IUPHAR/BPS Guide to PHARMACOLOGY in 2016: towards curated quantitative interactions between 1300 protein targets and 6000 ligands. Nucleic Acids Res 44:D1054-68 |
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| Catterall, William A; Swanson, Teresa M (2015) Structural Basis for Pharmacology of Voltage-Gated Sodium and Calcium Channels. Mol Pharmacol 88:141-50 |
| Chen-Izu, Ye; Shaw, Robin M; Pitt, Geoffrey S et al. (2015) Na+ channel function, regulation, structure, trafficking and sequestration. J Physiol 593:1347-60 |
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